Colorimetric determination of N-hydroxylated metabolites in microsomal studies

A colorimetric method is described which can be used for the routine determination of primary and secondary N-hydroxylamino compounds in drug metabolism studies using microsomal preparations. The assay is carried out on the supernatant obtained after protein precipitation of the incubation mixture. The method is based on the reduction of ferric ion by the hydroxylamino moiety with the resulting ferrous ion being quantitated by coupling with 2,4,6-tripyridyl-s-triazine to form a purple color with a maximum absorbance at 595 nm. The method is specific to primary and secondary hydroxylamino compounds and calibration curves can be obtained in the range of concentrations of 1 to 20 μg/ml. Hydroxamic acid, amido, amino, phenolic, nitro, nitroso, oxime, nitrone, aldehyde, and ketone compounds do not produce any color reaction when analyzed by the same method. The method is simple, rapid, and many samples can be analyzed in a short period of time.     

Effects of 3,5,3'-triiodothyroacetic acid (TRIAC) on protein metabolism of genetically obese or non-obese Zucker rats

Genetically obese female rats (fa/fa) and their lean littermates (Fa/-) were given oral administration of 3,5,3-triiodothyroacetic acid (TRIAC) (20 micrograms/ 100 g of body weight/ day) during 4 weeks. Metabolism of proteins was evaluated in several organs and in skeletal muscle after intraperitoneal injection of 14C and 3H-leucine 6 days and 16 hrs respectively before the sacrifice of animals. We have determined radioactivity of 14C and 3H and the 3H/14C ratio. No significant differences were found in lean and obese rats except in skeletal muscle. The relative protein turnover in skeletal muscle is significantly higher in the obese rats than in the lean rats. Treatment by TRIAC decreases the body weight gain in obese rats compared with controls but it has no statistically significant effect on the relative protein turnover in either obese or lean rats.     

In Vitro Release of Endogenous β- Alanine, GABA, and Glutamate, and Electrophysiological Effect of β-Alanine in Pigeon Optic Tectum

The efflux of 20 amino acids, induced by either high K+ concentration or veratrine, was determined in pigeon tectal slices. Ca2+-dependent, K+-induced release of beta-alanine, gamma-aminobutyric acid (GABA), and glutamate was observed. Veratrine caused release of the same amino acids plus glycine in a tetrodotoxin-sensitive manner. beta-Alanine had a strong inhibitory effect on the activity of tectal neurons which was blocked by strychnine but not by bicuculline. The results indicated a transmitter function for beta-alanine in the optic tectum, and were consistent with the previously proposed transmitter role of GABA and glutamate in this structure.     

Substitution of the flavin chromophore with lipophilic side chains: A novel membrane redox label

Summary The synthesis of amphiphilic flavins substituted with C₁₈-hydrocarbon sidechains in positions 3, 5, 7, 8 and 10 is described. 3-, 7-, and 10-amphiflavins were obtained by new total syntheses. Furthermore, 3-amphiflavin was obtained by C₁₈-alkylation of natural flavin in the oxidized state, whereas 5-amphi(dihydro)flavin was obtained by alkylation under reducing conditions.In the course of these studies, a novel, selective oxidation reaction was found taking place with the 8-methyl group of natural flavins. In this way lumiflavin and riboflavin derivatives could be converted directly to flavin-8-nor-8-carboxylic acids or the corresponding alkyl esters.The new flavin derivatives lend themselves for incorporation into lipid vesicles, thus yielding the basis for model studies of anisotropic flavin chemistry and redox transfer through membranes, as described in the concomitant paper (Schmidt, W., Hemmerich, P. 1981).J. Membrane Biol. 59:129. The new flavins are characterized by means of absorption, fluorescence, and proton nuclear magnetic resonance spectroscopy.     

Characteristics of synaptosomal tyrosine uptake in various brain regions: Effect of other amino acids

Tyrosine uptake by rat synaptosomes was maximal after 5–10 min of incubation and at 30°C; uptake was inhibited by dinitrophenol (10^?4 M) or ATP (10^?3 M) and increased by reducing sodium concentrations or increasing calcium or potassium. The best model for uptake is a two-carrier system, in which one carrier shows high-affinity uptake and the other may be diffusional. Both uptake mechanisms are more concentrated in catecholamine-rich brain areas, and are inhibited $\text{in vitro}$ by other large, neutral amino acids. At physiologic amino acid brain concentrations, each system probably carries about half of the tyrosine into the nerve terminal.     

Foliar flavonoid distribution during Spinacia chloroplast isolation

The flavonoid glycoside accumulation level was compared in class A and in class C spinach chloroplast suspensions. Class A chloroplasts (up to 97% intact) contained about 0.4% of the total flavonoid glycosides present in leaves. Further purification of 97%-intact chloroplast suspensions, through a Percoll gradient, reduced the glycoside level to less than 0.15%. On the other hand, class C chloroplasts (100% broken plastids) contained between 10 to 30 times more flavonoids than intact Percoll purified chloroplasts. These results indicate that chloroplasts could bind vacuolar glycosides during their period of isolation. This hypothesis was confirmed by controlled contamination experiments using a cell-free supernatant as a source of vacuolar glycosides. Furthermore, the level of flavonoids in chloroplasts could be decreased to a level close to that obtained in intact Percoll purified chloroplasts by washing with soluble polyvinylpyrolidone. The results presented in this paper demonstrate the importance of maintaining the physiological integrity of plastids during the course of organelle isolation when investigating flavonic compartmentation in leaves.     

Deoxyribonucleic acid relatedness inYersinia enterocolitica andYersinia enterocolitica-like organisms

Yersinia enterocolitica andY. enterocolitica-like strains were characterized by DNA relatedness. These strains formed four distinct DNA relatedness groups: (i) the 5 classical biotypes ofY. enterocolitica sensu stricto as designated by Wauters; (ii) strains that are rhamnose positive and also positive in tests for melibiose, α-methyl-d-glucoside, raffinose, and Simmons' citrate; (iii) strains that are rhamnose positive but negative in tests for melibiose, α-methyl-d-glucoside, and raffinose; (iv) sucrose-negative, Voges-Proskauer-negative, trehalose-positive strains.